Download Distribution of hepatitis C virus genotypes among intravenous drug

Turkish Journal of Medical Sciences
Turk J Med Sci
(2016) 46: 66-71
© TÜBİTAK
doi:10.3906/sag-1411-169
http://journals.tubitak.gov.tr/medical/
Research Article
Distribution of hepatitis C virus genotypes among intravenous drug users in the
Çukurova region of Turkey
1,*
2
1
3
4
Enver ÜÇBİLEK , Bahri ABAYLI , Mahmut Bakır KOYUNCU , Durdane MIDIKLI , Süveyda GÖZÜKÜÇÜK ,
5
6
1
1
Alper AKDAĞ , Osman ÖZDOĞAN , Engin ALTINTAŞ , Orhan SEZGİN
1
Department of Gastroenterology, Faculty of Medicine, Mersin University, Mersin, Turkey
2
Department of Gastroenterology, Çukurova Dr. Aşkım Tüfekçi State Hospital, Adana, Turkey
3
Department of Infectious Diseases, Adana State Hospital, Adana, Turkey
4
Department of Infectious Diseases, Dr. Ekrem Tok Psychiatry Hospital, Adana, Turkey
5
Department of Infectious Diseases, Ceyhan State Hospital, Adana, Turkey
6
Department of Gastroenterology, Tarsus State Hospital, Mersin, Turkey
Received: 01.12.2014
Accepted/Published Online: 07.03.2015
Final Version: 05.01.2016
Background/aim: The most common hepatitis C virus (HCV) genotype in Turkey is genotype 1. However, there has not been a study
about the distribution of HCV genotypes among intravenous drug users (IVDUs) in the Çukurova region of Turkey. This study was
planned to understand if there is a difference between IVDUs and the normal population.
Materials and methods: Between May 2010 and May 2014, anti-HCV positive IVDUs who applied to the 6 hospitals in the Çukurova
region of Turkey were included in this study. Their HCV genotypes were studied.
Results: Ninety-seven anti-HCV positive IVDUs were screened in terms of HCV RNA and genotype. Ten were excluded from the study
because their HCV RNA results were negative. Fifty-one of the 87 patients (58.6%) had genotype 3. Genotype 2 was detected in 26
(29.9%) and genotype 1 was detected in 10 (11.5%) patients.
Conclusion: HCV genotypes seem to be different between the normal population and IVDUs according to studies worldwide. Among
IVDUs, we detected a dominance of genotype 3 and genotype 2, which is apparently different from the normal population. The reason
for this difference can be simply explained by infection through shared needles. However, there may still be a different immunological
response in IVDUs, the investigation of which may lead to further studies.
Key words: Hepatitis C, HCV genotypes, intravenous drug users
1. Introduction
The hepatitis C virus (HCV) is a blood-borne virus and
an important cause of chronic liver disease, liver cirrhosis,
and hepatocellular carcinoma. It is also one of the most
prominent causes of liver transplantation, especially in
Western countries (1,2). Its worldwide prevalence is about
2%, which means that almost 180,000,000 individuals are
infected with HCV. However, among different countries
the prevalence shows extensive variation; for instance,
while its prevalence is 1.3%–1.6% in the United States, it
increases up to 15% in Egypt (3,4). In Turkey, the subject
of this study, the prevalence is around 0.4%–1% (5,6).
The highest risk groups for HCV infection consists
of intravenous drug users (IVDUs), patients who have a
history of blood transfusion, and hemodialysis patients,
since the virus is blood-borne (7). Due to the fact that
*Correspondence: [email protected]
66
with today’s technology blood transfusion can be done
with much safer methods, the percentage of transfusionrelated infections has considerably decreased. However,
unfortunately, intravenous drug usage is still one of the
most important causes of viral transmission due to shared
needles; this fact results in IVDUs being the highest risk
group for HCV infection (7). HCV prevalence among
IVDUs ranges from 30% to 90% worldwide (8–10).
The HCV genome includes 9400 nucleotides and
reveals a high genetic heterogeneity, which results in the
emergence of multiple genotypes (11). So far, 6 major
genotypes and multiple subtypes have been identified, the
distribution of which also varies in different geographical
regions (12,13). Genotype 1 is the dominant genotype in
North Europe, North America, South and East Europe,
and Japan, whereas genotype 2 is detected less frequently
ÜÇBİLEK et al. / Turk J Med Sci
in these regions. On the other hand, genotype 3 is most
common in Southeast Asia, genotype 4 in Egypt and
Middle Africa, and genotype 5 in South Africa (14–16).
In Turkey, studies show that genotype 1 (with subtype
1b) is the dominant type in the country, some studies even
giving the resulting prevalence as up to 100%. However
some recent studies indicate that, as of now, frequencies
of other genotypes are increasing (17–23). It is well known
that the HCV genotype directly affects the success of the
treatment and the clearance rate; therefore, the diagnosis
of the HCV genotype in the pretreatment phase is critical
for medication.
During the treatment and tracking of IVDU patients
in the Çukurova region, it was noticed that the frequency
of genotype 3 had increased significantly. Since there
have not been any studies about the distribution of HCV
genotypes among IVDUs in Turkey, this study was planned
to investigate if there is a difference between IVDU and
non-IVDU patients.
2. Materials and methods
The IVDUs who applied to gastroenterology and infectious
diseases clinics of the State Hospital of Adana Çukurova,
the Dr. Ekrem Tok Psychiatric Hospital of Adana, the
State Hospital of Adana, the State Hospital of Ceyhan, the
State Hospital of Tarsus, the hospital of Mersin University
Medical School, and the State Hospital of Kahramanmaraş
between May 2010 and May 2014 with an HCV antibody
positive diagnosis were examined and evaluated in terms
of HCV RNA levels and genotypes.
Serological anti-HCV, HBsAg, and anti-HIV tests
were done using the enzyme-linked immunosorbent assay
(ELISA) method (Abbott Laboratories, USA). HCV RNA
and hepatitis B virus (HBV) DNA levels were measured
by the real-time polymerase chain reaction (RT-PCR)
technique with the help of a Cobas TaqMan 48 kit (Roche
Diagnostics, USA). Determination of the HCV genotypes
was accomplished with another kit, an AMPLIQUALITY
HCV-TS (AB Analitica, Italy), which is also referred to as
the “line probe assay” (LiPA). The LiPA quantifies the HCV
genotypes by reverse hybridization of the amplification
products that are already measured by Cobas TaqMan RTPCR from the 5’UTR viral region. Special probes, made
up of the unique oligonucleotides of HCV genotypes, are
immobilized on nylon strips that contain these products.
The statistical analysis was executed by using the
Mann–Whitney U test and differences were assumed to be
significant at P < 0.05.
3. Results
Ninety-seven patients with positive HCV antibody
diagnosis were involved in this study. After evaluating
their HCV RNA levels and genotypes, 10 of them were
excluded from the group of interest because their HCV
RNA levels were negative. Of the remaining 87 patients, 2
of them were female (2%) and the other 85 patients were
male (98%) with an average age of 27.4 ± 6.8 years. HBV–
HCV coinfection was detected in only 1 patient and the
HBV DNA level of this patient was found to be negative.
None of the patients had HIV infection. While one of the
patients had an acute HCV infection, the rest were chronic
HCV patients.
After examining the genotype distribution, genotype
3 was identified in 51 (58.6%) of the 87 patients. Among
these, genotype 3a was detected as a subtype in 2 patients
while a subtype could not be assigned for the rest.
Genotype 2 was detected in 26 (29.9%) patients, with 1 of
them having genotype 2b, and genotype 1 was identified in
10 (11.5%) with 5 of them having genotype 1a.
HCV RNA levels of the patients ranged from 1.10 ×
102 IU/mL to 3.3 × 107 IU/mL, with an average HCV RNA
level of 1.28 × 106 ± 3.91 × 106 IU/mL. Mean HCV RNA
levels were 1.28 × 106 ± 1.66 × 106 IU/mL in genotype 1,
2.22 × 106 ± 6.67 × 106 IU/mL in genotype 2, 8.15 × 105
± 1.66 × 106 IU/mL in genotype 3. Although HCV RNA
levels were lower in genotype 3 patients, statistically there
was not any significant difference between these groups in
terms of this or binary HCV RNA comparisons (Table 1).
Table 1. Characteristics of the study population.
Genotype 1
Genotype 2
Genotype 3
P-value
Frequency
11.5%
29.9%
58.6%
NA*
Sex (male/female)
10 / 0
26 / 0
49 / 2
NA
Age (years)
27.8
25.8
28.1
0.38
HCV RNA (IU/mL)
1.28 × 106
2.22 × 106
8.15 × 105
0.36
*NA: Not available.
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ÜÇBİLEK et al. / Turk J Med Sci
4. Discussion
Identification of the HCV genotype is an important
parameter for the course and treatment of the disease.
Studies so far have clarified that genotype 1 is the most
frequent genotype in the Turkish population (17–23).
Our previous research, focusing only on the genotype
distribution in Mersin, is also consistent with this fact
as genotype 2 and 3 were detected with rates of 2% and
4%, respectively, whereas genotype 1 and subtype 1b were
diagnosed in 92% and 85% of the patients in the study
group (17). However, when IVDUs come into question,
the distribution changes completely, with genotype 3
(58.6%) in first place, followed by genotype 2 (29.9%), and
genotype 1 only identified in a few patients (11.5%). Other
genotypes were not detected in any of the patients.
Although the prevalence of genotype 1 is about 90% in
Turkey, some recent studies have shown that the frequency
of other genotypes is increasing in some regions of the
country. Genotype 4 in Kayseri, genotype 2 in Gaziantep,
and genotype 3 in İstanbul and Adana are being detected
much more often compared to older studies (18,22,24,25).
In a recent study from Adana and Antakya, genotype 1
turned out to be the most prevalent (59%), followed by
genotype 2 (14%) and genotype 3 (26%) (18). In the latter
study, it could not be clarified why these genotypes were
seen at such high rates. However, our study may explain
these unusual results since in the study of Öztürk et al. (18)
the route of transmission was not mentioned. Probably,
IVDU were included in that study and they may have led
to unexpectedly high rates.
Another study recently published reveals that there was
no change in the distribution of genotypes in the past 10
years in Turkey, with genotype 1 being the most frequent
genotype with 90% prevalence (26). Performed among 500
patients, the HCV genotypes and subtypes according to
the NS5B, E1, and 5’UTR regions were diagnosed at rates
of 92.5%, 93.5%, and 87.7% for genotype 1b and 6.7%,
5.6%, and 6.6% for genotype 1a, respectively, and also in all
3 analyses, genotypes 2 and 3 were detected at a rate of less
than 1%. Eventually, in this study it was deduced that the
phylogenetic analysis of NS5B and E1 regions indicated
more accurate and consistent data compared to 5’UTR,
because genotype 1 rates found with 5’UTR analysis are
lower than the other two; however the deviation is still
small. We used the 5’UTR viral region for determining
the HCV genotypes in our study based on the fact that
this method is being used in many regions of the world
(3,24,27).
HCV genotypes in IVDU patients were found to be
variable in different studies from various regions of the
world (3,7,27–36) (Table 2). For instance, studies from
China showed that the most prevalent genotype among
all patients was genotype 1, followed by genotype 2. On
the other hand, when focusing specifically on IVDUs,
Table 2. Genotype distribution among IVDU from various countries.
Study
HCV genotype (%)
1
2
3
4
5
6
China (24)
12
0
41
0
0
47
China (25)
11
0
35
0
0
46
China (26)
24
0
41
0
0
33
France (3)
52.9
2.3
36.8
7.5
0
0
Nigeria (29)
75
25
0
0
0
0
Greece (11)
22
2.4
65
10.4
0
0
Lebanon (33)
21.4
0
57.1
17.9
0
0
Belgium (27)
38
2
49
9
0
0
Iceland (28)
60
0
37.5
0
0
0
Cyprus (30)
43
0
57
0
0
0
Italy (31)
45.5
3
35
15
0
0
Slovakia (32)
41.3
0
57.7
0
0
0
This study
11.5
29.9
58.6
0
0
0
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ÜÇBİLEK et al. / Turk J Med Sci
genotype 6 becomes dominant, followed by genotype
3 (28–30). In Europe, for all patients genotype 1 is the
predominant genotype; however, an increase in genotype
3 frequency is noted among IVDUs according to some
studies (3,7,27,31,33–35). For another example, in
Lebanon, although the predominant genotype is genotype
4, genotype 3 was diagnosed at a high rate among IVDUs
(36) (Table 2).
In this study, we determined that the most common
genotypes among IVDUs are genotypes 3 and 2, whereas
genotype 1 was identified with a very low rate. These
findings are unexpected because even in geographically
close regions to Turkey, like Cyprus or Greece, genotype
1 has not been identified with such a low rate (7,33). In
addition, HIV is expected to be diagnosed at a high rate
among IVDUs; however, in our study none of the patients
had HIV infection (3,37). Lastly, HBV–HCV coinfection
was determined only in 1 patient, whose HBV DNA was
detected as negative. Another implication of the study
is that IV drug usage among women is not common in
Turkey compared to other countries, since only 2% of the
patients were female in our study (38–42).
This research had some limitation due to the
characteristics of the subject of interest. First, in IVDUs,
adherence to HCV treatment is not at the desired level.
A great number of patients cannot complete the therapy
due to restarting IV drug usage or the side effects of the
therapy (7). Besides, a lack of information about the history
and duration of drug usage and also of the records of the
progress in the treatment limits further investigation and
therefore a variety of conclusions can be drawn. In addition,
there is also a restricted amount of data about the effects of
different HCV genotypes on transmission and replication
of the disease as the studies are focused on the effects
of genotype on the treatment. For example, especially
intermittent viremia can be observed by reinfection in
IVDU, affecting further consecutive transmissions. Also,
for some cases, the viruses that could initially escape from
the immune response may develop rebound viremia. It is
suspected that some human monoclonal antibodies may
have a neutralizing effect; however, unfortunately, the
correlation with the genotypes has not yet been clarified
(43).
To sum up, it is not clear why the prevalence of HCV
genotypes among IVDUs is different than in normal
patients. This situation can be related to infection
through sharing needles; however, that is far too simple
an explanation for such a significant difference. Also, if
that were the case, the IVDUs who were infected with
genotype 1, the dominant genotype all over the country,
would also spread the disease through needle sharing and
cause an increase in genotype 1 percentage. Therefore, it
seems likely that different immunologic reactions catalyze
the fast spread or help the persistence of genotype 3 and 2
HCV RNA in IVDUs. This is not within the scope of this
study but, nevertheless, it can be an inspiration for further
research.
In conclusion, the prevalence of HCV genotypes
among IVDUs is different from the normal population
all over the world. It was found that rare genotypes, like
genotype 3 and genotype 2, are seen commonly among
IVDUs in the Çukurova region of Turkey. This study is
important because this is the first of its kind in Turkey and
it needs to be supported by new studies from other regions
of the country.
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